Department of Tissue Culture and Stem Cells

The main topics studied in the Department are isolation, labelling and the use of stem cells for the treatment of brain injury, spinal cord and neurodegenerative diseases. Various types of cells (mesenchymal stem cells, neural precursor cell lines derived from fetal spinal cord, or from induced pluripotent cells) are studied, together with anti-inflammatory substances for their potential to promote the regeneration of nervous tissue. Macroporous polymeric hydrogels are used as suitable carriers for cell growth in in vitro cultures as well as for in vivo implantations facilitating the regeneration of the injured tissue. The aim of the cell therapy is to repair, replace or improve biological functions of the damaged neural tissue. For in vivo imaging of grafted cells and drug delivery we utilize magnetic nanoparticles, which are characterized in terms of cytotoxicity and genotoxicity and their influence on grafted cells and host tissue.

Important results in 2015

Mesenchymal stem cells reduce the working memory deficit in Alzheimer's disease model.Stem cell transplantation may have a positive influence and slow the progression of some neurodegenerative diseases. In our study, we transplanted human mesenchymal stem cells (MSCs) into the lateral ventricle of 8 months old transgenic mice (AD-3xTg), which mimic the symptoms of Alzheimer's disease (AD). We studied the changes in the spatial reference and working memory, and the effect of transplanted MSCs on neurogenesis in the subventricular zone (SVZ). We also monitored the levels of harmful oligomer amyloid 56kDa (Ab*56), and the amount of the enzyme glutamine synthetase (GS), which is important for regulating the levels and metabolism of glutamate in the brain, in entorhinal and prefrontal cortex and in the hippocampus, i.e. in the structures that are related with cognitive functions. In 14 months old mice treated with MSC we observed preserved working memory, which may be a result of preserved levels of GS and significantly reduced levels of Ab*56 in the entorhinal cortex (Figure 1). These changes, observed six months after transplantation, were also accompanied by increased cell proliferation in the SVZ. Since the transplanted cells survive in the body of the recipient only for a limited period of time, it is likely that the observed effects could be even more pronounced in case of repeated administration of the stem cells at regular intervals during the life spam of the 3xTg mice.

Working memory test in mice with AD. The red line represents the second trial to find the island in the water maze. The ability to remember the position of the islet is shown at the top left of the chart. AD mice had in the entorhinal cortex reduced level of harmful amyloid oligomer Aβ*56. The graph on the bottom right. AD - Alzheimer's disease, EC - entorhinal cortex, ctrl - control animals were age-matched.

Important results in 2014

1. Neural precursors from induced pluripotent cells (iPS-NP) significantly improve motor function in rats with spinal cord injury.Grafted cells (iPS-NP) survived well and slowly mature into different neuronal phenotypes (GABAergic, Serotonergic and motoneurons). In addition, they produced growth factors, stimulating neuronal sprouting. As a result, animals with spinal cord injury significantly improved their motor function. Rats were able to support their body weight and perform stepping, so they scored well in tests requiring movement coordination, such as beam walking.

2. Mesenchymal stem cells increase lifespan of animals with amyotrofic lateral sclerosis (ALS).We studied the effect of human mesenchymal stem cells (MSCs) in the treatment of an experimental model of ALS. We found that application of MSCs improved motor performance and muscle strength and led to an extended lifespan. MSCs partially rescued motor neuron (MN) loss and decreased apoptosis. MSCs transplantation is therefore a safe procedure able to promote CNS remodeling and regeneration.

The effect of MSC application. After the appearance of fi rst disease symptoms SOD1 rats were treated (arrow) with MSCs (intrathecally, 5×105 cells/50μl) or vehicle-injected (DMEM, 50μl). Shortly after delivery of MSCs disease progression has been slowed down shown by the digger muscle strength (A) and higher motor activity (B). MSC-treated rats lived signifi cantly longer (C, D) and preserved higher number of ventral motor neurons (E) compared with vehicle-injected littermates. We found that SOD1 rats have deteriorated perineuronal nets structure around motor neurons and that application of MSCs partially preserved their structure (F).

2. Modification of polymeric hydrogels based on methacrylate, serving to bridge lesions and as carriers for cell therapy.In cooperation with the Institute of Macromolecular Chemistry ASCR, we developed and tested polymer hydrogels based on methacrylate with different types of pores and surface modified positively charged peptides or RGD sequences. Hydrogels were implanted into a model of acute spinal cord hemisection. The results showed that the positive charge and network pore structure of the RGD sequences on the surface promotes cell growth, both in vitro and in vivo, as well as the ingrowth of axons and blood vessels of the host.

Methacrylate hydrogel with a modified surface bridged the spinal cord lesion (A) and promoted ingrowth of axons (B), blood vessels (C) and astrocytes (D). Served as a suitable carrier for transplanted stem cells.